1. Field
The invention is in the field of cytocentrifugation devices used to centrifugally deposit sediment material, such as cells, from a sediment containing sample onto a microscope slide, and particularly in the field of sample chambers for receiving the sample and applying the sample to the microscope slide in such devices.
2. State of the Art
Cytocentrifugation devices are small centrifuges particularly designed for centrifuging small samples of body fluids, such as blood, to separate cells or other sediment materials therefrom and deposit the cells or other sediment materials onto a microscope slide for further examination and evaluation. Cytocentrifugation devices generally include a rotor to which a plurality of sample receiving chambers in association with microscope slides are removably secured. The fluid sample containing the sediment material to be examined is placed into a sample receiving well in the sample chamber. During centrifugation, the sample flows by centrifugal force against the microscope slide so that cells or other sediment materials in the sample are forced against the microscope slide and generally adhere thereto. During centrifugation, the sample is held in a sedimentation chamber against the microscope slide which forms an end of the sedimentation chamber. The sedimentation chamber may serve as the sample receiving well, or the sample receiving well may be separated from the sedimentation chamber and arranged so that the sample flows from the sample receiving well into the sedimentation chamber upon the start of centrifugation. After the centrifugation, the sample receiving chamber is removed from the cytocentrifugation device and the microscope slide is also removed from the cytocentrifugation device for examination and evaluation. Depending upon the cytocentrifugation device, the sample chamber and microscope slide may be removed from the cytocentrifugation device as a unit with the microscope slide later separated from the sample chamber for examination and evaluation, or the two can be removed from the cytocentrifugation device separately. However, in either case, it is desirable that the liquid portion of the sample be removed from contact with the microscope slide at the termination of centrifugation so that the liquid does not interfere with the sediment deposited on the slide and so that the slide and sample chamber can be removed from the cytocentrifuge device or otherwise separated from one another without the mess and possible contamination that might result from the liquid running uncontrollably out of the sedimentation chamber.
Several methods have been used to remove the liquid portion of the sample from around the microscope slide either during centrifugation or when centrifugation stops. In some sample chambers, a removal passage is provided leading from the opening of the sedimentation chamber against the microscope slide to a collection system. A vacuum source is connected to the removal passage to suck the sample liquid from the sedimentation chamber after centrifugation stops. Examples of such sample chambers are shown in U.S. Pat. Nos. 4,306,514, 4,314,523, 4,327,661, 4,423,699, and 4,428,323. These systems all require the complication of providing vacuum lines connected to arms of the centrifuge rotor, and the use of a vacuum waste collection system.
Systems having liquid removal passages have been developed where absorbent material is positioned in the removal passage to eliminate the need for the vacuum source. The sample liquid is drawn from the sedimentation chamber-microscope slide interface by the absorbent material in the removal passage. Examples of these systems are shown in U.S. Pat. Nos. 4,574,729 and 4,576,110. A problem with systems using absorbent material in a removal passage to absorb the liquid portion of the sample is that the absorption of the sample by the absorbent material begins before the centrifugation of the sample is completed. This means that a portion of the sample with sediment material is lost in the removal passage before such sediment can be deposited on the slide. This reduces the number of cells or the amount of other sediment material that can be deposited on the slide reducing the effectiveness of the separation of the sediment materials from the liquid portion of the sample.
Many cytocentrifuge systems currently in use position absorbent material, such as filter paper or filter card, between the end of the sedimentation chamber and the microscope slide around the periphery of the sedimentation chamber opening to allow and control flow of sample liquid into the filter paper or card so that the sample liquid is slowly but completely absorbed by the filter paper or filter card by the end of centrifugation. Examples of these systems are shown in U.S. Pat. Nos. 4,391,710, 4,678,579, 5,252,228, 5,380,435, and 5,466,371. These systems are designed to be used with small samples of material to be evaluated. Thus, such systems normally have sample chambers to hold about 300 to 500 microliter samples, with some holding up to about one milliliter of sample. In some situations, it is desirable to use larger samples, up to about twelve milliliters, and usually up to about six milliliters. This is the case where it is desirable to have more cells from a sample for observation than are available in a small sample of less than about 1 milliliter. However, with large volume samples, it is impractical to absorb all of the sample liquid at the sample chamber-microscope slide interface under centrifugal force. With large volume samples, it is necessary to provide a seal between the sample chamber and the microscope slide so that sample fluid does not escape during centrifugation and to provide for removal of the sample liquid after centrifugation is completed. While the vacuum removal system referred to earlier can be used in such circumstances, a simpler system is preferred.
An approach to removal of the sample liquid from the sedimentation chamber-microscope slide interface when centrifugation is completed is to make the sedimentation chamber slope downwardly away from the microscope slide so that the sample is held in the lower portion of the sedimentation chamber when centrifugation is not taking place. However, the sloping of the chamber downwardly so that the top and bottom sedimentation chamber walls are not aligned with the centrifugal force applied to the sample, has been found to result in uneven distribution of the sediment on the microscope slide giving a concentration of cells at the bottom of the sample area and weak cell densities at the top of the sample area. A further problem in using larger sample volumes and larger sample chambers is that the cells or other sediment materials tend to settle in the sample chamber prior to centrifugation and are applied unevenly to the microscope slide during centrifugation. U.S. Pat. Nos. 5,470,758 and 5,589,400 address this problem by providing a sample chamber for a large sample, wherein the sample receiving well, which also serves as the sedimentation chamber to hold the sample during centrifugation and application to the microscope slide, is not only sloped downwardly, but is also provided with a plurality of baffle plates therein which define a tortuous path for sample flowing into the sample receiving well as the sample receiving well is filled. The baffle plates collect some of the sediment in the sample fluid on the baffle plates as the sample fills the sample receiving well and sedimentation chamber prior to centrifugation. The baffle plates spread the sediment throughout the chamber rather than allowing it all to settle in the bottom of the chamber.
As indicated, in the device of U.S. Pat. Nos. 5,470,758 and 5,589,400, the sedimentation chamber is slopped downwardly so that the sample placed in the sedimentation chamber remains in the lower part of the chamber away from the microscope slide until centrifugation takes place. At that time, the sample chamber rotates slightly, but the chamber still remains at a sloped orientation to the microscope slide. The sample is forced to the upper end of the sedimentation chamber and against the microscope slide by the centrifugal force of the centrifugation. At the end of centrifugation, the sample liquid decants and flows by gravity back into the lower portion of the sedimentation chamber and away from the slide. The use of the baffles in the sedimentation chamber to spread the sediment material has not solved the problem of uneven distribution of the sample on the slide. It has been found in use of such baffled chamber that rather than an even disposition of sediment on the slide, the sediment is applied to the microscope slide in striations corresponding to the baffle locations. This is in addition to the weaker cell densities at the top of the sample area due to the sloping chamber. In addition, with the sample retained in the lower portion of the sedimentation chamber, avoiding spillage of the decanted fluid during chamber removal requires some care in handling. Further, the fluid does not always decant completely and droplets can remain on the microscope slide where the chamber contacts the slide surface primarily at the bottom of the collection area. When the chamber is pulled away from the slide, these droplets can flow down on the slide and contaminate the rotor. Also, the residual fluid can cause smearing of the sample and cause cells to dislodge and migrate out of the collection area. Cells may also be lost during fixation as a result of the migration.
A sample chamber for large samples that provides a more even distribution of the sediment on the slide as well as better drainage and removal of the sample liquid left after centrifugation is needed.